Staphylococci are hardy and ubiquitous colonizers of human skin and mucous membranes and were among the first human pathogens identified. These bacteria constitute a medically important genera of microbes as they are known to produce two types of disease, invasive and toxigenic.
Invasive infections are characterized generally by abscess formation affecting both skin surfaces and deep tissues. In addition, Staphylococcus aureus (S. aureus) is the second leading cause of bacteremia in cancer patients. Osteomyelitis, septic arthritis, septic thrombophlebitis and acute bacterial endocarditis are also relatively common.
There are also at least three clinical conditions resulting from the toxigenic properties of Staphylococci. The manifestation of these diseases result from the actions of exotoxins as opposed to tissue invasion and bacteremia. These conditions include: Staphylococcal food poisoning, scalded skin syndrome and toxic shock syndrome.
S. aureus are non-motile, non-sporulating gram-positive cocci 0.5-1.5 μm in diameter, that occur singly and in pairs, short chains, and irregular three-dimensional clusters. S. aureus can grow over a wide range of environmental conditions, but they grow best at temperatures between 30° C. and 37° C. and at a neutral pH. They are resistant to desiccation and to chemical disinfection, and they tolerate NaCl concentrations up to 12%. However, growth of S. aureus becomes unusually sensitive to a high-NaCl concentration by decreasing the Ca2+ concentration in growth media allowing for autolysis (Ishikawa, Microbiology and Immunology, 2000: 44(2):97-104).
Humans constitute the major reservoir of the S. aureus bacteria. The cross-sectional carriage rate in adults is 15 to 40 percent. The mucous membranes of the anterior nasopharynx are the principal site of carriage. Other sites include the axillae, the vagina, the perineum and occasionally the gastrointestinal tract. Colonization by S. aureus may be intermittent or persistent and is probably influenced by both microbial and host factors as well as by the nature of the competing non-Staphylococcal flora.
The frequency of S. aureus infections has risen dramatically in the past 20 years. This has been attributed to two main factors. The first factor is an increasing population of people with weakened immune systems. The second factor has been the emergence of multiple antibiotic resistant strains. It is no longer uncommon to isolate S. aureus strains which are resistant to some or all of the standard antibiotics. Active efflux of various toxic compounds from the cell to the outer medium is a universal mechanism that bacteria have evolved to protect themselves against the adverse effects of their environments. Antibiotics are expelled from the cells by membrane transporter proteins called multidrug resistance (MDRs) pumps. The NorA protein of S. aureus is an MDR pump that mediates the active efflux of hydrophilic fluoroquinolones from the cell (Ubukata, et al. Antimicrob. Agents Chemother., 1989: 33(9):1535-9), conferring low-level resistance upon the organism. NorA is also capable of transporting additional structurally diverse compounds, indicating that it has a broad substrate specificity. NorR, a protein that shares 60% similarity with B. subtilis MarR and S. aureus SarA regulatory proteins, may function to repress the transcription of NorA (Truong-Bolduc, et al., ASM 2001 General Meeting, May 22, 2001: Abstract A-56).
The rise in the frequency of S. aureus infections has created a demand for both new anti-microbial agents and diagnostic tests for this organism. Accordingly, there is a need for better understanding of factors which regulate infectivity and growth of S. aureus. Genes identified as involved in the infectivity and/or growth of S. aureus include the arlS regulator, involved in adhesion (Fournier and Hooper, Journal of Bacteriology, 2000: 182(14):3955-64), the pbpC gene, which affects the rate of autolysis (Pinho et al., Journal of Bacteriology, 2000: 182(4):1074-9), the atl gene, which encodes a protein having amidase and N-acetylglucosaminidase activity (Sugai, et al., Journal of Bacteriology, 1997:179:2958-2962), lytN (Sugai, Journal of Infection and Chemotherapy, 1997:3:113-127), lytRS (Brunskill and Bayles, Journal of Bacteriology, 1996: 178(19):5810-2), lrgA and lrgB (Fujimoto et al., Journal of Bacteriology, 2000: 182(17):4822-8), and lytM identified in autolysis-deficient mutants of S. aureus (Ramadurai and Jayaswal, Journal of Bacteriology, 1997: 179(11):3625-31).
A new polypeptide of S. aureus which regulates autolytic processes has now been identified. The nucleic acid sequences encoding this polypeptide, referred to herein as Rat, regulator of autolytic activity, which regulates autolytic processes has been cloned and sequenced. Further, it has been shown that mutations in the nucleic acid sequences encoding Rat render S. aureus more susceptible to lysis by antibiotics.